ABSTRACT: Metastatic melanoma is one of the fastest growing tumor types in the US, one of the top 5 cancers among Veterans, and is also one of the most challenging malignancies to treat. Currently immune therapies targeting the check-point inhibitors such as CTLA-4 and/or PD-1 can enhance the T cell response to tumor an these inhibitors have shown great success and as a result have moved to first line standard of care for most metastatic melanoma patients. Ipilimumab (targeting CTLA-4) combined with nivolumab (targeting PD-1) has yielded a 59% response rate in melanoma patients. Melanoma patients with an increased mutational load, and those with NF1 mutation, respond better to anti-PD1 or anti-PDL1 therapy. However, side effects are often extensive, producing severe adverse events in >60% of patients including myocarditis for combined anti-CTLA4 and anti-PD1 therapy. Clearly, refinements of current treatments are needed to enhance survival of metastatic melanoma patients. Approximately 40% of melanoma tumors have mutation or loss of CDKN2A. One of the genes encoded by CDKN2A, INK4a is important for inhibiting the cell cycle kinases, CDK4/6. This results in loss of restrictions on cell cycle and enhanced proliferation of melanoma tumor cells. We have shown that inhibitors of the cell cycle kinases, CDK4/6, can both slow the growth of some melanoma tumors and also enhance T cell recruitment into the tumors. Moreover, approximately 80% of melanoma patients have activating mutations in the RAS/RAF/MEK/MAPK pathway (30% with mutation in NRAS and 51% with mutation of BRAF). Another 23% have alterations resulting in activation of the PI3K pathway. Recently rigosertib (RGS), a non-ATP competitive multi-kinase inhibitor, has been shown to inhibit the RAS/RAF/MEK and PI3K signaling pathways which are also crucial for melanoma tumor growth. We propose here to determine the effectiveness of combining CDK4/6 inhibitors with RGS in comparison to anti-PD1 therapy which is standard of care. We hypothesize that RGS, which targets multiple kinases, will be effective for treating RAS mutant melanoma in combination with CDK4/6 inhibitors. We also propose that an even greater response will occur when check point inhibitors such as anti- PD1 or with activating antibody to CD137 are added to the RGS and CDK4/6i combined therapy regime. In this research grant we will examine the ability of CDK4/6 inhibitor combined with RGS to inhibit the growth of melanoma in immune competent mouse models of melanoma, including NRAS mutant, BRAF mutant, and B16 melanoma (WT for NRAS and BRAF). We will also examine the effects of CDK4/6 inhibition combined with RGS on the immune response to the tumor and characterize the effectiveness of combining RGS with anti-PD1 or other check point inhibitors, or with addition of anti-CD137 treatment in immune competent mouse models of melanoma. Finally, we will evaluate the effectiveness of combining CDK4/6 inhibitor and RGS with anti-PD1 or anti-CD137 for treatment of melanoma patient derived xenografts using humanized mouse models. This study should lead to new and improved therapeutic options for treating our Veterans with metastatic melanoma.